Increase of metallothionein-immunopositive chloride cells in the gills of brown trout and rainbow trout after exposure to sewage treatment plant effluents

Metallothionein, a biomarker of exposure and toxicity of heavy metals, has been detected in the gills of brown trout (Salmo trutta fario L.) and rainbow trout (Oncorhynchus mykiss Richardson) by means of immunohistochemistry. A very prominent labelling of chloride cells was found after exposure to diluted sewage plant effluents. No significant increase was observed in either the number of labelled cells or their labelling intensity after exposure to water of a polluted river compared to fish kept in tap water. These results do not correlate with findings of a histopathological study, suggesting that the metal levels at the sewage treatment plant were too low to produce gross histopathology. A comparison between the species indicated that the rainbow trout showed a generally higher metallothionein expression than the brown trout

Immobilization of Shewanella oneidensis MR-1 in diffusive gradients in thin films for determining metal bioavailability

Assessing metal bioavailability in soil is important in modelling the effects of metal toxicity on the surrounding ecosystem. Current methods based on diffusive gradient thin films (DGTs) and Gel-Integrated Microelectrode are limited in their availability and sensitivity. To address this, S. oneidensis, an anaerobic iron reducing bacterium, was incorporated into a thin layer of agarose to replace the polyacrylamide gel that is normally present in DGT to form biologically mobilizing DGT (BMDGT). Viability analysis revealed that 16-35% of the cells remained viable within the BMDGTs depending on the culturing conditions over a 20 h period with/without metals. Deployment of BMDGTs in standardized metal solutions showed significant differences to cell free BMDGTs when cells grown in Luria Broth (LB) were incorporated into BMDGTs and deployed under anaerobic conditions. Deployment of these BMDGTs in hematite revealed no significant differences between BMDGTs and BMDGTs containing heat killed cells. Whether heat killed cells retain the ability to affect bioavailability is uncertain. This is the first study to investigate how a microorganism that was incorporated into a DGT device such as the metal reducing bacteria, S. oneidensis, may affect the mobility of metals

Effects of Altering Freshwater Chemistry on Physiological Responses of Rainbow Trout to Silver Exposure

The influence of different water Cl- (50–600 μM), Ca2+ (50–1,500 μM), Na+ (50–1,500 μM), or dissolved organic carbon (DOC, 0.31–5 mg/L) levels on silver-induced physiological and biochemical perturbations of rainbow trout were investigated. Fish were acclimated to soft water (50 μM; Cl-, Ca2+, and Na+), then exposed to 3.7 μg/L Ag (as AgNO3) for 6 h, which resulted in a reduction in Na+ influx from the water, an inhibition of gill sodium- and potassium-activated adenosine triphosphatase (Na+/K+-ATPase) activity, and an accumulation of silver on the gills. Increasing the water Cl- or DOC levels ameliorated the silver toxicity. However, increasing water Ca2+ or Na+ concentration did not reduce the silver-induced physiological and biochemical perturbations. The free silver ion (Ag+) concentrations (calculated from MINEQL+, a geochemical speciation computer program) showed a negative correlation with the Na+ influx rates and gill Na+/K+-ATPase activity. However, gill silver levels did not correlate to Ag+ concentrations and no correlation was found between gill silver levels and either Na+ influx rates or gill Na+/K+-ATPase activity. These results support the notion that the [Ag+] concentration is of major importance when assessing silver toxicity in fish, and that this should be taken into account in regulatory strategies for silver in the natural environment

Liver transcriptome analysis of Atlantic cod (Gadus morhua) exposed to PCB 153 indicates effects on cell cycle regulation and lipid metabolism

Background: Polychlorinated biphenyls (PCBs) are persistent organic pollutants (POPs) with harmful effects in animals and humans. Although PCB 153 is one of the most abundant among PCBs detected in animal tissues, its mechanism of toxicity is not well understood. Only few studies have been conducted to explore genes and pathways affected by PCB 153 by using high throughput transcriptomics approaches. To obtain better insights into toxicity mechanisms, we treated juvenile Atlantic cod (Gadus morhua) with PCB 153 (0.5, 2 and 8 mg/kg body weight) for 2 weeks and performed gene expression analysis in the liver using oligonucleotide arrays. Results: Whole-genome gene expression analysis detected about 160 differentially regulated genes. Functional enrichment, interactome, network and gene set enrichment analysis of the differentially regulated genes suggested that pathways associated with cell cycle, lipid metabolism, immune response, apoptosis and stress response were among the top significantly enriched. Particularly, genes coding for proteins in DNA replication/cell cycle pathways and enzymes of lipid biosynthesis were up-regulated suggesting increased cell proliferation and lipogenesis, respectively. Conclusions: PCB 153 appears to activate cell proliferation and lipogenic genes in cod liver. Transcriptional up-regulation of marker genes for lipid biosynthesis resembles lipogenic effects previously reported for persistent organic pollutants (POPs) and other environmental chemicals. Our results provide new insights into mechanisms of PCB 153 induced toxicity.Peer ReviewedpublishedVersio

Safety and efficacy of furfuryl and furan derivatives belonging to chemical group 14 when used as flavourings for all animal species and categories

Following a request from the European Commission, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety and efficacy of 18 compounds belonging to chemical group 14 (furfuryl and furan derivatives with and without additional side-chain substituents and heteroatoms). They are currently authorised as flavours in food. This opinion concerns 13 compounds from this group. The FEEDAP Panel concludes that all compounds except 5-methylfurfural are safe at the proposed maximum use level for all animal species: furfural and furfuryl alcohol at 5 mg/kg complete feed; methyl 2-furoate and furfuryl acetate at 0.5 mg/kg complete feed; bis-(2-methyl-3-furyl) disulfide, furanmethanethiol, S-furfuryl acetothioate, difurfuryl disulfide, methyl furfuryl sulfide, 2-methylfuran-3-thiol, methyl furfuryl disulfide and methyl 2-methyl-3-furyl disulfide at 0.05 mg/kg complete feed. 5-Methylfurfural is safe at the proposed use level of 0.5 mg/kg complete feed for cattle, salmonids and non-food producing animals and at the use level of 0.3 mg/kg complete feed for pigs and poultry. No safety concern would arise for the consumer from the use of these compounds up to the highest safe level in feeds. Hazards for skin and eye contact and respiratory exposure are recognised for the majority of the compounds under application. Most are classified as irritating to the respiratory system. The concentrations considered safe for the target species are unlikely to have detrimental effects on the terrestrial and freshwater environments. Since all the compounds under assessment are used in food as flavourings, and their function in feed is essentially the same as that in food, no further demonstration of efficacy is necessary. In the absence of data on the stability in water for drinking, the FEEDAP Panel is unable to conclude on the safety or efficacy of the substances under this mode of delivery

Acute health risks related to the presence of cyanogenic glycosides in raw apricot kernels and products derived from raw apricot kernels

Amygdalin is the major cyanogenic glycoside present in apricot kernels and is degraded to cyanide by chewing or grinding. Cyanide is of high acute toxicity in humans. The lethal dose is reported to be 0.5–3.5 mg/kg body weight (bw). An acute reference dose (ARfD) of 20 μg/kg bw was derived from an exposure of 0.105 mg/kg bw associated with a non-toxic blood cyanide level of 20 micro mol (μM), and applying an uncertainty factor of 1.5 to account for toxicokinetic and of 3.16 to account for toxicodynamic inter-individual differences. In the absence of consumption data and thus using highest intakes of kernels promoted (10 and 60 kernels/day for the general population and cancer patients, respectively), exposures exceeded the ARfD 17–413 and 3–71 times in toddlers and adults, respectively. The estimated maximum quantity of apricot kernels (or raw apricot material) that can be consumed without exceeding the ARfD is 0.06 and 0.37 g in toddlers and adults, respectively. Thus the ARfD would be exceeded already by consumption of one small kernel in toddlers, while adults could consume three small kernels. However, consumption of less than half of a large kernel could already exceed the ARfD in adults

Regulated mitochondrial DNA replication during oocyte maturation is essential for successful porcine embryonic development.

Cellular ATP is mainly generated through mitochondrial oxidative phosphorylation, which is dependent on mitochondrial DNA (mtDNA). We have previously demonstrated the importance of oocyte mtDNA for porcine and human fertilization. However, the role of nuclear-encoded mitochondrial replication factors during oocyte and embryo development is not yet understood. We have analyzed two key factors, mitochondrial transcription factor A (TFAM) and polymerase gamma (POLG), to determine their role in oocyte and early embryo development. Competent and incompetent oocytes, as determined by brilliant cresyl blue (BCB) dye, were assessed intermittently during the maturation process for TFAM and POLG mRNA using real-time RT-PCR, for TFAM and POLG protein using immunocytochemistry, and for mtDNA copy number using real-time PCR. Analysis was also carried out following treatment of maturing oocytes with the mtDNA replication inhibitor, 2',3'-dideoxycytidine (ddC). Following in vitro fertilization, preimplantation embryos were also analyzed. Despite increased levels of TFAM and POLG mRNA and protein at the four-cell stage, no increase in mtDNA copy number was observed in early preimplantation development. To compensate for this, mtDNA appeared to be replicated during oocyte maturation. However, significant differences in nuclear-encoded regulatory protein expression were observed between BCB(+) and BCB(-) oocytes and between untreated oocytes and those treated with ddC. These changes resulted in delayed mtDNA replication, which correlated to reduced fertilization and embryonic development. We therefore conclude that adherence to the regulation of the timing of mtDNA replication during oocyte maturation is essential for successful embryonic development

Safety and efficacy of manganese compounds (E5) as feed additives for all animal species: manganous carbonate; manganous chloride, tetrahydrate; manganous oxide; manganous sulphate, monohydrate; manganese chelate of amino acids, hydrate; manganese chelate of glycine, hydrate, based on a dossier submitted by FEFANA asbl

The Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) has assessed five manganese compounds: manganous chloride, tetrahydrate; manganous sulphate, monohydrate; manganous oxide; manganese chelate of amino acids, hydrate; manganese chelate of glycine, hydrate. The safety assessment is based on the assumption that the current maximum total contents of manganese authorised in feed are respected. All manganese compounds under application are considered safe for all animal species/categories. The mean manganese intake of the European population includes already the manganese from animal products and does not pose a toxicological concern. The effect of dietary manganese fed to animals on tissue concentrations is limited. Supplementation of feed with the manganese compounds under assessment would consequently not affect consumer exposure and is of no concern for consumer safety. All manganese compounds are considered as eye irritants, the manganese chelate of glycine and the manganese chelate of amino acids as irritants to skin and the latter one as dermal sensitiser. However, the presence of nickel in all additives may induce contact dermatitis. Exposure to manganese in dust of all additives and to nickel (except manganous chloride and manganous oxide) poses a risk to users by inhalation. The use of the manganese compounds under assessment in animal nutrition for all animal species is not expected to pose a risk to the environment. The manganese compounds under assessment are recognised as efficacious sources of manganese in meeting animals’ requirements

Presence of microplastics and nanoplastics in food, with particular focus on seafood

Following a request from the German Federal Institute for Risk Assessment (BfR), the EFSA Panel for Contaminants in the Food Chain was asked to deliver a statement on the presence of microplastics and nanoplastics in food, with particular focus on seafood. Primary microplastics are plastics originally manufactured to be that size, while secondary microplastics originate from fragmentation. Nanoplastics can originate from engineered material or can be produced during fragmentation of microplastic debris. Microplastics range from 0.1 to 5,000 μm and nanoplastics from approximately 1 to 100 nm (0.001–0.1 μm). There is no legislation for microplastics and nanoplastics as contaminants in food. Methods are available for identification and quantification of microplastics in food, including seafood. Occurrence data are limited. In contrast to microplastics no methods or occurrence data in food are available for nanoplastics. Microplastics can contain on average 4% of additives and the plastics can adsorb contaminants. Both additives and contaminants can be of organic as well of inorganic nature. Based on a conservative estimate the presence of microplastics in seafood would have a small effect on the overall exposure to additives or contaminants. Toxicity and toxicokinetic data are lacking for both microplastics and nanoplastics for a human risk assessment. It is recommended that analytical methods should be further developed for microplastics and developed for nanoplastics and standardised, in order to assess their presence, identity and to quantify their amount in food. Furthermore, quality assurance should be in place and demonstrated. For microplastics and nanoplastics, occurrence data in food, including effects of food processing, in particular, for the smaller sized particles (< 150 μm) should be generated. Research on the toxicokinetics and toxicity, including studies on local effects in the gastrointestinal (GI) tract, are needed as is research on the degradation of microplastics and potential formation of nanoplastics in the human GI tract

Safety and efficacy of pyridine and pyrrole derivatives belonging to chemical group 28 when used as flavourings for all animal species

Following a request from the European Commission, the EFSA Panel on Additives and Products or Substances used in Animal Feed (FEEDAP) was asked to deliver a scientific opinion on the safety and efficacy of nine compounds belonging to chemical group 28 (pyridine, pyrrole and quinoline derivatives). They are currently authorised as flavours in food. The FEEDAP Panel concludes that piperine, 3-methylindole, indole, 2-acetylpyridine and 2-acetylpyrrole are safe at the proposed maximum use level of 0.5 mg/kg complete feed for all animal species; trimethyloxazole, 3-ethylpyridine, pyrrolidine and 2,6-dimethylpyridine are safe at the proposed use level of 0.5 mg/kg complete feed for cattle, salmonids and non-food-producing animals, and at the use level of 0.3 mg/kg complete feed for pigs and poultry. No safety concern would arise for the consumer from the use of these compounds up to the highest safe level in feeds. Hazards for skin and eye contact, and respiratory exposure are recognised for the majority of the compounds under application. Most are classified as irritating to the respiratory system. The concentrations considered safe for the target species are unlikely to have detrimental effects on the terrestrial and fresh water environments. As all the compounds under assessment are used in food as flavourings, and their function in feed is essentially the same as that in food, no further demonstration of efficacy is necessary. In the absence of data on the stability in water for drinking, the FEEDAP Panel is unable to conclude on the safety or efficacy of the substances under this mode of delivery